The Role of Bacteriosin as Food Preservative Mahendra Pal*, Azeb Gebretensay*, Tilaye Shiberu*, Mukarim Abdurahman, Olga Karanfil** Department of Microbiology, Immunology and Public Health, College of Veterinary Medicine, Addis Ababa University,P.B.No.34,Debre Zeit, Ethiopia ** Ethiopian Meat and Dairy Industry Development Institute,P.B.1573, Debre Zeit, Ethiopia Email :
[email protected]
ABSTRACT Food safety is a growing public health concern of both in the developing, and developed nations. In the developing countries, food borne disease often goes unreported, as strong surveillance, and reporting systems are unavailable. However, the high incidence of diarrhoeal diseases suggests the major underlying food safety problems. Food borne diseases are prevalent all over the world, and are responsible for high morbidity, and mortality. It is estimated that food borne infections causes 76 million cases, 300,000 hospitalizations, and 500 deaths annually in the United States. The safe food supply is the best way to reduce the incidences of food borne diseases. The consumption of chemically preserved foods raises the concern among the consumer for a naturally produced antimicrobial agents.Bacteriocins are antibacterial proteins produced by many lactic acid bacteria in fermented, and non-fermented foods. Presently, nisin is the only bacteriocin which is used as a food preservative in many countries of the world. Bacteriocins are inhibitory against food borne pathogens, and bacteriocin resistance is not usually genetically determined. The simultaneous application of bacteriocins, and non-thermal processing technologies to improve the shelf life of foods is attractive since foods produced using these nonthermal technologies usually have better sensory, and nutritional qualities compared with products produced using conventional thermal processing methods. As bacteriocins are very effective, and safe, their uses in the food industries are recommended. Moreover, the safety of other bacteriocins with their potential application in various types of food should be assessed. Key words: Bacteriocin, Consumer, Food, Nicin, Public health, Preservative, Safety
INTRODUCTION Nature has given many types of foods to mankind. Everybody expects that the food they eat is wholesome, and safe for consumption. The greatest threat to quality, and safety of our food comes from the microbial spoilage (Pal, 2013). The food spoilage is wasteful, and costly; and can adversely affect the economy, and erode the confidence This paper is dedicated to the memory of Marie Curie ,the First women Scientist ,who discovered “ Radium” , and won two Nobel Prize, one for Physics in 1903, and second for Chemistry in 1911.
of the consumers. It is well established that food is a valuable source of nutrients for certain microbes. As they grow on the food, they may cause problems such as bad taste, unpleasant smell, and poor appearance. More importantly, the growth of microbes may lead to dangerous levels of toxins in the food. This makes the food unfit to be eaten by the people, and hence it leads to food scarcity (Pal, 2013). Preservatives are a type of food additive which are added to food to prolong shelf life, and keep the products from being broken down by microorganisms (Pal, 2014). Moulds, bacteria, and yeasts can cause food spoilage, and are found practically everywhere including the air we breathe (Pal, 2013). Most preservatives today are actually fungistatic in their action. That means they prevent the growth of fungi, moulds and yeasts. They have little effect on bacteria but using a combination of preservatives, with antibacterial properties can give good protection. Food preservatives help to control the spread of bacteria which can cause life threatening illnesses such as salmonellosis or botulism. Preservatives are commonly used in these foods such as low fat spreads, cheeses, margarine, mayonnaise and dressings, bakery products, and dried fruit preparations (Pal, 2014). Among bio-preservatives, bacteriocin has caught the attention of food scientists to be used as a natural food biopreservative due to its antimicrobial activity against food spoilage, and pathogenic bacteria. Different bacteriocin producing strains of lactic acid bacteria as well as Bacillus spp. have been isolated for this purpose but the keen interest towards bacteriocin of lactic acid bacteria wordwide is due to their essential role in majority of food fermentation, flavor development and preservation of food products along with proving safer for health. Preservation action of lactic acid bacteria is due to production of lactic acid, acetic acid, hydrogen peroxide as well as bacteriocin resulting from metabolic activity of organism (Cleveland et al., 2002). Microbiologists around the world got interested in bacteriocin producing microorganism to overcome this problem that fulfill the requirement of food preservation. Although many other type of bacteriocin such as subtilin, cerein, thuricin, plantaricin etc., have been isolated and characterized, and are still in a process of getting commercial status to be used as food preservatives so far only one bacteriocin, Nisin has been given the status of preservative to be added in food items commercially (Ogunbanwo et al., 2003).
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The use of microorganisms in food fermentation is one of the oldest methods for producing and preserving food. Much of the world depends upon various fermented foods that are staples in the diet. Till the end of 1950, a very few of the food items were processed, and packaged. The processed and packaged foods were luxury item in colonial times but after 1960 these food items were in great demand globally due to growing urbanization, breakdown of large families into nuclear families, and increase in the number of working women (Sharma et al., 2006). Many methods of food preservation are used to keep safe. Processes such as freezing, canning, pickling and drying are attempted to remove one or more of the factors necessary for the growth of food spoiling microbes (Cleveland et al., 2002). The present communication highlights the growing significance of bacteriocin as food preservative. FOOD PRESERVATIVES Chemical preservatives help to delay the growth of microbes in food. If it were not for food preservatives, our bread would go mouldy too fast, and our salad dressing would turn even faster.Traditional methods of preserving with salt, refrigeration, pickling, and freezing are not always optimal for foods that need to be kept fresh. For this reason, food preservatives are a vital additive to the daily foods we eat (Pal, 2014). Food preservatives can be classified into two general categories: antimicrobials or antioxidants. Antimicrobials inhibit the growth of yeasts, bacteria, and molds. Antioxidants slow air oxidation of lipids and fats, which causes foods to go rancid too quickly. Aside from simply giving food a longer shelf life, food preservatives are vital to preserving food during shipping. This is how people in Maine are able to enjoy California raisins, or for olive oil from Spain to be used in Missouri salad dressing (Pal, 2014). There is a wide variety of food preservatives used by cooks, and food manufacturers to extend shelf life and improve storage of both cooked and raw foods. Many natural preservatives, like salt and sugar, are often used in preserving canned fruits, processed meats as well as various types of canned or jarred vegetables. Nitrates and nitrites are common artificial preservatives found in many processed meats, including lunch meat and bacon, and are made from naturally occurring sodium chloride found in table salt. Antioxidants also play a role in preserving fresh and processed foods, as do proper at home preservation methods like freezing, pickling or jarring foods (Pal, 2014). Food manufacturers use an array of food preservatives to keep food fresh, and stable, as well as to extend shelf life, and prevent natural decay (Pal,2013).One of the most common food preservatives is salt, suitable for many uses such as preserving meat, fish, canned vegetables and fruits, as well as frozen desserts, and vegetables. Salt is one of the most popular food preservatives used at home for preserving raw meat, as salt helps to draw out moisture from the meat through a process called osmosis. This is often called curing the meat, and is one of the oldest food preservation methods known (Pal, 2014). If there would been no preservatives, one could not enjoy many of the great foods the world has to offer. The common antimicrobial preservatives include sodium nitrate, calcium propionate, disodium EDTA, potassium sorbate, sodium benzoate, and sulfites; and all are FDA approved (Pal, 2014).
BACTERIOCINS Bacteriocins are produced by bacteria, and possess antibiotic properties, but bacteriocins are normally not termed antibiotics in order to avoid confusion, and concern with therapeutic antibiotics that can potentially illicit allergic reactions in humans (Cleveland et al., 2001).Most of the bacteriocins are bactericidal with some exceptions Leucocin A UAL 187 being bacteriostatic. Inhibitory activity of bacteriocin producing strains are mostly confined to Grampositive bacteria. Bacteriocins are bactericidal to sensitive cells.The sensitivity of Gram-positive, and sensitivity of Gram- negative bacteria towards bacteriocins has been demonstrated on the basis of cell wall composition. It has been observed that Gram-negative bacteria become sensitive towards bacteriocin action, if it is destabilized by physical or chemical stresses (Ray et al., 2001). Gramnegative bacteria possess an additional layer so called outer layer which is composed of phospholipids proteins, and lipopolysacharides, and this layer is impermeable to most molecules. The presence of porin in this layer allows free diffusion of molecular with a molecular mass below 600 Da. The smallest bacteriocin produced LAB are approximately 3 kD, and too large to reach their target cytoplasmic membrane (Abee et al., 1995). It has been proposed that nisin acts on the cytoplasmic membrane of Gram- positive bacteria to cause lesions (Bower et al., 1995). Following nisin treatment, whole or intact sensitive cells, and membrane vesicles exhibit efflux of amino acid, and cations. The loss of these substances depletes proton motive force, which ultimately interferes with cellular biosynthesis. These events result in collapse of membrane potential, and ultimately cause cellular death (Nettles and Barefoot, 1993). Similarly, other bacteriocins such as Lactococin A, Pediocin J D, etc., have also been reported to cause dissipation of the membrane potential, and increase in membrane permeability to ions leading to collapse of proton motive force (Bruno and Montville, 1993). Recent interest to isolate bacteriocin producing strains is due to its effectiveness against food spoilage/pathogenic bacteria, and also due to its proteinaceous nature which made it safer for human consumption. It is assumed to be degraded by protease in the gastrointestinal track (Cleveland et al., 2002). The digestive enzymes rapidly inactivate bacteriocin and consequently it cannot alter bacterial microflora in the intestinal track. Many regulatory agencies have advocated the use of hurdle concept, i.e. combining several physical and chemical methods for preservation in sub optimal levels to control microbial growth. Pediocin when combined with low dose irradiation showed a greater inhibitory effect on growth of L. mesenteroides. Till today, many food scientists have advanced packaging techniques to incorporate antimicrobial agents into food products to control microbial growth, and enhance safety, and shelf life of food products (Appendini and Hotchkiss, 2002). Bacteriosin as Food Preservative Effect of pH on Bacteriocin Production Influence of pH on bacteriocin production has also been studied. Optimum pH for bacteriocin production is between 6.0 to 7.0 (Skytta et al., 1993). Some of the bacterial strains were found to produce active bacteriocin at pH range 5.8 to 7. 9 .While strain Leuconostoc MF215B was found to produce bacteriocin at pH 6.0 but not at pH 7.5 (Blom et al.,
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1999). Strain L. gelidin showed best production at 6.5. Similarly, amylovorin L471 produced at pH 6.5.While C. piscicola produced bacteriocin at pH 7.0 (Herbin et al., 1997). None of the investigators so far reported bacteriocin production in the alkaline range, i.e. between pH 9.0 and 12.0. So it could be concluded that optimum bacteriocin production is exhibited between the range of pH 5.0 and 8.0, and production declined beyond these limits. Effect of Temperature on Bacteriocin Production Incubation temperature for obtaining high yield of bacteriocin also varies from strain to strain. Most of the strains which have been isolated so far showed bacteriocin production at temperature 30°C to 37°C. However, strain D53 isolated from the vegetables has been found to produce bacteriocin from 10°C to 37°C (Uhlman et al., 1992). Brevibacterium linens produces bacteriocin at 25°C than at 30°C, and no significant growth or production was observed at 37°C. Similarly, high amount of bacteriocin of L. sake was produced at 25–30°C while decline in production was observed at 33.5°C followed by nil production at 34.5°C or higher (Diep et al., 2000). Bacteriocin production of L. plantarumY21 was obtained at 30°C. In milk, bacteriocin was produced during incubation at 37°C or under temperature gradient reproducing at first 24 h of soft cheese manufacture (Tarelli et al., 1994). Several studies confirmed that the best temperature range for bacteriocin production is 30°C to 37°C. The combination of two different bacteriocin can also be used to increase shelf life of food e.g. two bacteriocins named nisin, and pediocin PA-1/ACH were used in combination to prevent spoilage in dairy, meat and fish foods. Instead of using combination of two bacteriocins, some workers have successfully combined nisin with lysozyme or nisin with some lactate against food spoilage bacteria. Bacteriocin preparation can be used at 1–2% level, and bacteriocin activity should kept below 5000 AU/g or ml of food (Gravesen et al., 2002). More advanced genetic methods may provide an excellent advantage for improving shelf life of the cheese. Inhibition of spoilage, pathogenic bacteria was attained by introducing bacteriocin genes into cheese starter culture by conjugation (Ross and Hill, 1998). In one study, hot dogs were inoculated with two bacteriocin based on preservatives BPI i.e, (mixture of Pediocin ACH and nisin A) and BP2 i.e, (BP1+bacterial metabolites (organic acid), and then challenged before vacuum packaging with two common processed meat spoilage bacteria, L. mesenteroides and L. viridesans. During 9 weeks of storage at refrigeration temperature, both bacterial strains grew beyond spoilage detection levels I in the control samples but not in BP treated samples. In other studies, viability and growth of three Gram-negative pathogens especially spores of C. botulinum in beef products at 25°C were reduced by BP preparation. Similar studies were also conducted by preparing processed meat products using BP as an ingredient in the raw material and inoculating the finished products with spoilage bacteria L. mesenteroides. During storage, the growth of L. monocytogenes was controlled in the product prepared with bacteriocin preparation. These results clearly indicated that bacteriocin based products have the potential to control spoilage and pathogenic bacteria and ensure safety to enhance shelf life of food products (Ray et al., 2001). The effect of bacteriocin in comparison to chemical
preservative has been studied in milk, cheese, apple juice. Bacteriocin of L. brevis, B. mycoidess showed encouraging results in all treated food items (Sharma and Gautam, 2007). In one of the study immersion solutions of enterocin AS-48 alone or in combination with chemical preservatives was applied on fresh green asparagus, alfalfa, and soyabean sprouts to inhibit growth of L. monocytogenes CECT 4032 (Mollinos et al., 2005).The combination of nisin (25ìg/ml) with 1% H 2O2, 1% sodium lactate, and 0.5% citric acid, and was found to reduce the contamination of L. monocytogenes, and Escherichia. coli. This mixture has been recommended as a washing treatment to decontaminate melon surfaces and hence to improve the microbial safety, and quality of fresh cut melon (Ukuku et al., 2005). Though bacteriocins play a major role in food industry, these preparations have been also applied in sugar processing, seed treatment, antibacterial cream, cosmetics, mouth wash, and toothpaste (Gravesen et al., 2002). CONCLUSION It may be concluded from the study of various authors that, that day is not far off when chemical preservative will be replaced by bio-preservative completely or partially in the food ultimately providing safer,and healthier food to the consumer, and leading to a revolution in the food industry. Although many bacteriocins have been isolated, and characterized, only a few have demonstrated commercial potential in food application. At this time, nisin is the only purified bacteriocin approved for food use in the U.S. It has been used as a food preservative in more than 50 countries, mainly in cheese, canned vegetables, various pasteurized dairy, liquid egg products, and salad dressings. The applications of other bacteriocins in food preservation have been studied intensively. The use of pediocin PA-1 for food biopreservation has been commercially exploited, and is covered by several U.S. and European patents. Fermentate containing pediocin PA-1, AltaTM, is commercially available, and used as a food preservative to increase shelf life, and inhibit the growth of bacteria, especially L. monocytogenes in ready-to-eat meats. Lacticin 3147, which is active over a wider pH range than nisin, is expected to find applications in non-acid foods. Since bacteriocins for use as food preservatives have relatively narrow activity spectra, and are generally not active against Gram- negative bacteria, it can be expected that nisin, and other bacteriocins will continue to be incorporated, and developed into hurdle concept technologies for food preservation. ACKNOWLEDGEMENT The authors are very grateful to Prof.R.K.Narayan for going through our manuscript. REFERENCES Abee, T., Krockel, L. and Hill, C. (1995): Bacteriocin: Modes of action and potential in food poisoning. International Journal Food Microbiology 28:169–185. Appendini, P. and Hotchkiss J.H. (2002): Review of antimicrobial food packaging. Innovative Food Science Emerging Technology 3:113–126. Blom, H., Katla, T., Holck, A., Sletten, K., Axelsson, L. and Holo, H. (1999): Characterization, production and purification of leucocin H, a two-peptide bacteriocin from leuconostoc MF 215 B. Current Microbiology 39: 43–48. Bower, C.K., Mcgure, J. and Deschel, M.A. (1995): Influence on the antimicrobial activity of surface adsorbed nisin. Journal Industrial Microbiology 15:227–233. Bruno, M.E.C. and Montville, T.J. (1993): Common mechanistic action of bacteriocin from lactic acid bacteria. Applied Environmental Biotechnology 40:143–150. Cleveland, J., Chiknids, M. and Montiville, T.J. (2002): Multimethod assessment of commercial nisin preparations. Journal of Industrial Microbiology and Biotechnology 29:228–232.
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used as stabilizers in many processed foods. Animals exposed to aluminum in the womb and during development show neurological effects such as changes in behavior, learning and motor response. Neurotoxicity has occurred in people undergoing dialysis who received large intravenous doses of unpurified water, but a direct link between aluminum food additives and neurological effects has not been proven (Schreeder 1983; EFSA 2008b). A link with Alzheimer’s disease and other neurodegenerative disorders has been proposed, but the association remains unclear (Bondy 2013). While significant scientific uncertainty remains around whether there may be links between aluminum-based food additives and health effects, their widespread use warrants putting them on the “watch list.” What you should do Read food labels to identify aluminum-based additives and consider alternatives. Use EWG’s Food Scores to find foods without aluminum additives. Contd. from Page 30
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